Chemical switches for detecting reactive chemical agents

Abstract

Certain embodiments of the present invention relate to the preparation of microbeads that exhibit a “turn on” fluorescence response within seconds of exposure to an analyte vapor (e.g., a chemical warfare agent or a reactive stimulant). This sensing approach is modeled after the mechanism for inhibition of acetylcholinesterase enzyme activity, and utilizes a specific and irreversible reaction between phosphonyl halides and a fluorescent indicator. The present invention also relates to a sensor and a method for sensing an analyte through detection of changes in the fluorescing properties of the inventive microbeads.

We claim: 
     
       1. A coated microbead, comprising a microbead and a coating, wherein said coating comprises a fluorescent compound represented by I or a salt thereof: 
       
         
           
           
               
               
           
         
         wherein, independently for each occurrence, W is O, S, or NR A ; Y is O, S, or NR A ; Z is O or NR A ; R A  is hydrogen, alkyl, acyl, or aralkyl; R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 11 , and R 12  are independently selected from the group consisting of hydrogen, halide, pseudohalide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, carbamoyl, hydroxyl, alkoxy, aryloxy, aralkoxy, acyloxy, thio, sulfonyl, sulfhydryl, sulfamoyl, sulfonamido, sulfate, sulfonate, sulfoxido, seleno, nitro, amino, imino, amido, phosphonate, phosphinate, silyl, trifluoroalkyl, cyano, and —(CH 2 ) m R B ; R B  is hydrogen, halide, pseudohalide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, carbamoyl, hydroxyl, alkoxy, aryloxy, aralkoxy, acyloxy, thio, sulfonyl, sulfhydryl, sulfamoyl, sulfonamido, sulfate, sulfonate, sulfoxido, seleno, nitro, amino, imino, amido, phosphonate, phosphinate, silyl, trifluoroalkyl, or cyano; and m is 1-10 inclusive; 
         provided that at least one of R 1 , R 2 , R 3  and R 4  is 
       
       
         
           
           
               
               
           
         
       
       wherein M is O; E 2  is hydroxyl, alkoxy, aralkoxy, sulfhydryl, amino, trifluoroalkyl, —O(CH 2 ) n R B , —NH(CH 2 ) n R B , or —S(CH 2 ) n R B ; E 3  is alkyl, cycloalkyl, heterocycloalkyl, aralkyl, heteroaralkyl, hydroxyl, alkoxy, aralkoxy, sulfhydryl, amino, or trifluoroalkyl; and n is 0-10 inclusive. 
     
     
       2. The coated microbead of  claim 1 , wherein Y is O; and Z is O. 
     
     
       3. The coated microbead of  claim 1 , wherein W is O. 
     
     
       4. The coated microbead of  claim 1 , wherein Y is O; Z is O; R 1  is H; R 2  is 
       
         
           
           
               
               
           
         
       
       R 3  is H; and R 4  is H. 
     
     
       5. The coated microbead of  claim 1 , wherein Y is O; Z is O; and W is O. 
     
     
       6. The coated microbead of  claim 1 , wherein Y is O; Z is O; W is O; R 5  is H; R 6  is H; R 7  is OH; R 8  is H; R 9  is H; R 11  is H; and R 12  is H. 
     
     
       7. The coated microbead of  claim 1 , wherein R 1  is H; R 2  is 
       
         
           
           
               
               
           
         
       
       R 3  is H; R 4  is H; and W is O. 
     
     
       8. The coated microbead of  claim 1 , wherein R 1  is H; R 2  is 
       
         
           
           
               
               
           
         
       
       R 3  is H; R 4  is H; W is O; R 5  is H; R 6  is H; R 7  is OH; R 8  is H; R 9  is H; is H; and R 12  is H. 
     
     
       9. The coated microbead of  claim 1 , wherein Y is O; Z is O; R 2  is 
       
         
           
           
               
               
           
         
       
       and R 7  is OH. 
     
     
       10. The coated microbead of  claim 1 , wherein Y is O; Z is O; R 1  is H; R 2  is 
       
         
           
           
               
               
           
         
       
       R 3  is H; R 4  is H; W is O; R 5  is H; R 6  is H; R 7  is OH; R 8  is H; R 9  is H; R 11  is H; and R 12  is H. 
     
     
       11. The coated microbead of  claim 1 , wherein said microbead comprises poly(2-vinylpyridine). 
     
     
       12. The coated microbead of  claim 1 , wherein said microbead comprises poly(2-vinylpyridine) and polystyrene. 
     
     
       13. The coated microbead of  claim 1 , wherein said microbead comprises poly(2-vinylpyridine) and a polystyrene functionalized with carboxylate moieties. 
     
     
       14. The coated microbead of  claim 1 , wherein said microbead comprises a polystyrene coated with poly(2-vinylpyridine), and said polystyrene is functionalized with carboxylate moieties. 
     
     
       15. The coated microbead of  claim 1 , wherein E 2  is alkoxy, amino, —O(CH 2 ) n R B , —NH(CH 2 ) n R B , or —S(CH 2 ) n R B . 
     
     
       16. The coated microbead of  claim 1 , wherein E 2  is ethoxy. 
     
     
       17. The coated microbead of  claim 1 , wherein E 3  is alkyl, cycloalkyl, or alkoxy. 
     
     
       18. The coated microbead of  claim 1 , wherein E 3  is methyl, ethyl, propyl, isopropyl, butyl, isobutyl, pentyl, isopentyl, 1,2,2-trimethylpropyl, cyclopentyl, cyclohexyl, or cycloheptyl. 
     
     
       19. The coated microbead of  claim 1 , wherein E 3  is methyl. 
     
     
       20. A method of detecting an analyte, comprising the steps of:
 contacting an indicator with a sample comprising an analyte, thereby providing a fluorescent compound; 
 exposing the fluorescent compound to light at a first wavelength; 
 measuring the fluorescence of the fluorescent compound at a second wavelength; and 
 comparing the fluorescence of the fluorescent compound at the second wavelength to the fluorescence of the indicator at the second wavelength, 
 wherein 
 the analyte is a compound having structure C: 
 
       
         
           
           
               
               
           
         
         wherein, independently for each occurrence, 
         M is O; 
         E 1  is F, Cl, Br, or I; 
         E 2  is hydroxyl, alkoxy, aralkoxy, sulfhydryl, amino, trifluoroalkyl, —O(CH 2 ) n R B , —NH(CH 2 ) n R B , or —S(CH 2 ) n R B ; 
         E 3  is alkyl, cycloalkyl, heterocycloalkyl, aralkyl, heteroaralkyl, hydroxyl, alkoxy, aralkoxy, sulfhydryl, amino, or trifluoroalkyl; 
         n is 0-10 inclusive; 
         R B  is hydrogen, halide, pseudohalide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, carbamoyl, hydroxyl, alkoxy, aryloxy, aralkoxy, acyloxy, thio, sulfonyl, sulfhydryl, sulfamoyl, sulfonamido, sulfate, sulfonate, sulfoxido, seleno, nitro, amino, imino, amido, phosphonate, phosphinate, silyl, trifluoroalkyl, or cyano; 
         the fluorescent compound is represented by structure I or a salt thereof: 
       
       
         
           
           
               
               
           
         
         wherein, independently for each occurrence, 
         W is O, S, or NR A ; 
         Y is O, S, or NR A ; 
         Z is O or NR A ; 
         R A  is hydrogen, alkyl, acyl, or aralkyl; 
         R 1 , R 2 , R 3 , R 4 , R 5 , R 6 , R 7 , R 8 , R 9 , R 11 , and R 12  are independently selected from the group consisting of hydrogen, halide, pseudohalide, alkyl, aralkyl, alkenyl, alkynyl, cycloalkyl, heterocyclyl, aryl, heteroaryl, aralkyl, heteroaralkyl, carboxyl, acyl, carbamoyl, hydroxyl, alkoxy, aryloxy, aralkoxy, acyloxy, thio, sulfonyl, sulfhydryl, sulfamoyl, sulfonamido, sulfate, sulfonate, sulfoxido, seleno, nitro, amino, imino, amido, phosphonate, phosphinate, silyl, trifluoroalkyl, cyano, and —(CH 2 ) m R B ; and 
         m is 1-10 inclusive; 
         provided that at least one of R 1 , R 2 , R 3  and R 4  is